The use of antibodies for treating human diseases is well established and has become more sophisticated with the introduction of genetic engineering. Several techniques have been developed to improve the utility of antibodies. These include: (1) the generation of monoclonal antibodies by cell fusion to create “hybridomas”, or by molecular cloning of antibody heavy (H) and light (L) chains from antibody-producing cells; (2) the conjugation of other molecules to antibodies to deliver them to preferred sites in vivo, e.g., radioisotopes, toxic drugs, protein toxins, and cytokines; (3) the manipulation of antibody effector functions to enhance or diminish biological activity; (4) the joining of other proteins such as toxins and cytokines with antibodies at the genetic level to produce antibody-based fusion proteins; and (5) the joining of one or more sets of antibody combining regions at the genetic level to produce bi-specific antibodies.
Proteins can be joined together through either chemical or genetic manipulation using methods known in the art. See, for example, Gillies et al., Proc. Natl. Acad. Sci. USA 89:1428–1432 (1992); and U.S. Pat. No. 5,650,150.
However, the utility of recombinantly-produced antibody-based fusion proteins may be limited by their rapid in vivo clearance from the circulation. Antibody-cytokine fusion proteins, for example, have been shown to have a significantly lower in vivo circulating half-life than the free antibody. When testing a variety of antibody-cytokine fission proteins, Gillies et al. reported that all of the fusion proteins tested had an a phase (distribution phase) half-life of less than 1.5 hours. Indeed, most of the antibody-based fusion proteins were cleared to 10% of the serum concentration of the free antibody by two hours. See, Gillies et al., BIOCONJ. CHEM. 4: 230–235 (1993). More recently, it was shown that antibody-based fusion proteins with reduced binding affinity for an Fc receptor have enhanced circulating half-lives. It was also shown that a reduced binding affinity for the Fc receptor interfered with some of the antibody effector functions such as antibody-dependent cellular cytotoxicity (ADCC), but did not interfere with other functions such as complement fixation or antigen binding. See Gillies at al., Cancer Res. 59(9):2159–66 (1999).
In some cases, such as the treatment of cancer or viral diseases, it would be desirable to maintain antibody effector functions and long circulating half-life. Therefore, there is a need in the art for additional methods of enhancing the in vivo circulating half-life of antibody-based fusion proteins.